doi 10.4067/s0718-19572012000300007 article morphological … · 2013-07-30 · key words:...

439Vol. 47, Nº3, 2012Revista de Biología Marina y Oceanografía

Revista de Biología Marina y OceanografíaVol. 47, Nº3: 439-450, diciembre 2012DOI 10.4067/S0718-19572012000300007Article

Morphological and molecular description of a newrecord of Graneledone (Cephalopoda, Octopodidae)

in the southeastern Pacific OceanDescripción morfológica y molecular de un nuevo registro de Graneledone (Cephalopoda, Octopodidae)

en el Océano Pacífico suroriental

Christian M. Ibáñez1, M. Cecilia Pardo-Gandarillas1, Elie Poulin1 and Javier Sellanes2,3

1Instituto de Ecología y Biodiversidad, Departamento de Ciencias Ecológicas, Facultad de Ciencias, Universidad de Chile, LasPalmeras 3425, Ñuñoa, Santiago, Chile. [email protected] de Biología Marina, Facultad de Ciencias del Mar, Universidad Católica del Norte, Larrondo 1281, Coquimbo,Chile3Centro de Investigación Oceanográfica en el Pacífico Sur-Oriental (COPAS), Universidad de Concepción, Casilla 160-C,Concepción, Chile

Resumen.- Los pulpos del género Graneledone habitan en aguas profundas y constituyen 8 especies reconocidas. Se

realizaron análisis filogenéticos de 4 especies de Graneledone con 2 marcadores moleculares (16S y COI), y se informa

sobre un nuevo registro de Graneledone para el Océano Pacífico frente a la zona centro-sur de Chile. Se obtuvieron 4

ejemplares de Graneledone sp. entre 436 y 1482 m de profundidad. Estos pulpos son de tamaño medio, no tienen saco de

tinta y tienen una sola fila de ventosas en sus brazos, que son de igual longitud. Se caracterizan por tener de 43 a 45

ventosas en el brazo hectocotilizado (tercero derecho); 6 a 7 laminillas por demibranquia; órgano del sifón en forma VV y

de 5 a 7 pliegues transversales en la lígula. Algunos rasgos morfológicos muestran una gran variación en comparación con

las especies del Pacífico, Atlántico y la Antártida. Los análisis filogenéticos moleculares apoyan la hipótesis de monofilia

de Graneledone.

Palabras clave: Graneledone, pulpos de aguas profundas, zona batial, Chile

Abstract.- Octopuses of the genus Graneledone inhabit in the deep-sea and are composed of 8 recognized species. Here we

conduct phylogenetics analyses of 4 species of Graneledone using 2 molecular markers (16S and COI), and report a new

record of Graneledone for the Pacific Ocean off south-central Chile. Four specimens of Graneledone sp. were collected from

436 to 1482 m depth. These octopuses are medium sized, have no ink sac and bear only one row of suckers in their arms,

which are of similar length. They are characterized by having 43 to 45 suckers in the hectocotylized arm (third right); 6 to

7 lamellae per demibranch; VV-shaped funnel organ and 5 to 7 transverse creases in the ligulae. Some morphological traits

show great variation compared with species from Pacific, Atlantic and Antarctic oceans. Molecular phylogenetic analysis

supports the hypothesis of monophyly of Graneledone.

Key words: Graneledone, deep-sea octopuses, bathyal zone, Chile

INTRODUCTION

Octopuses of the genus Graneledone Joubin, 1898generally live in the bathyal to abyssal zones rangingfrom 90 m to 2755 m depth and have been often reportedoccurring at particular ecosystems like hydrothermalvents and cold seeps (Voight 2000a, b, Guerrero-Kommritz2000, 2006). Graneledone is composed of 8 recognizedspecies (Norman & Hochberg 2005) found in all theoceans: G. antarctica Voss, 1976 and G. macrotyla Voss,1976 from the Antarctica, G. verrucosa (Verril, 1881) fromthe north Atlantic Ocean, G. gonzalezi Guerra, González& Cherel 2000 from the Kerguelen Island, G. yamanaGuerrero-Kommritz, 2000 from the southwest Atlantic, G.

boreopacifica Nesis, 1982 from the north Pacific and G.taniwha O’Shea, 1999 and G. challengeri (Berry, 1916)from the south Pacific Ocean. The validity of certainspecies and sub-species is still controversial, as they havebeen classified based on highly variable morphologicalcharacters, making differentiating them very difficult. Thisgenus is defined by having a body covered with warts,uniserial suckers, funnel organ VV shaped, no ink sac,crop reduced or absent, small gills, small posterior salivaryglands, homodont radula, and in males terminal organcoiled in spiral and large ligulae in hectocotylus (O’Shea1999, Guerrero-Kommritz 2000, Allcock et al. 2003). These

440 Ibáñez et al.New record of Graneledone for the SE Pacific

character states are derived among Octopodidaesuggesting the hypothesis of monophyly of Graneledone(Voight 1997, 2000a). Molecular divergence time estimationsuggests that Graneledone originated in the southernocean and their radiation into deep-sea was facilitated bythe thermohaline circulation (Strugnell et al. 2008a).However, this molecular analysis only included 3 species:G. antarctica, G. verrucosa and G. boreopacifica.

Until now, Pareledone charcoti Joubin, 1905,Pareledone turqueti Joubin, 1905, Thaumeledonerotunda (Hoyle, 1885) and Graneledone antarctica werethe only octopuses with uniserial suckers reported fromChilean waters (18°S-56°S, Thore 1959, Rocha 1997, Vegaet al. 2001, Vega 2009). However, recently specimens ofGraneledone have been collected off south-central Chile,one of them in the vicinity of a bathyal methane seep sitenear Concepción (~37°S, Ibáñez et al. 2011). This sitenamed Concepción Methane Seep Area (CMSA), hasproven to be an abundance hotspot for bothchemosymbiotic and heterotrophic fauna, and more thana hundred of megabenthic species have been recordedassociated with it (Sellanes et al. 2004, 2008). Among thisfauna, mollusks are one of the most important groups,with about 30 species present. In addition to commoncephalopods (sepiolids and squids), unidentified incirrateoctopod species belonging to Muusoctopus Gleadall 2004(formerly Benthoctopus) and Graneledone genus occurit the CMSA and adjacent areas (Ibáñez et al. 2006, 2009,2011).

In this study, we combine published data with newmitochondrial sequences to propose a phylogenetichypothesis of the deep-sea octopuses of the genusGraneledone. Moreover, we describe a new record ofGraneledone, comparing the type specimens bothmorphologically and genetically with their counterpartsfrom the Pacific, Atlantic and Antarctic Oceans to furtherelucidate their taxonomic position.

MATERIALS AND METHODS

The present work was based on 4 Graneledonespecimens captured off south-central Chile between 35°Sand 38°S (Fig. 1). Octopuses of this genus are the onlyones with uniserial suckers that have been reported sofar for the Chilean margin (see Ibáñez et al. 2011). One ofthe specimens was collected by trawling during a cruisededicated to the study of methane seep sites (29°S-45°S).The remaining 3 specimens were obtained as by-catchfrom commercial fishing activities, one from the long-line

fisheries targeting the Patagonian toothfish (Dissostichuseleginoides Smith, 1898), and the other 2 from shrimptrawling. Tissue samples were fixed in 96% ethanol formolecular analysis and the whole animal in 10% seawaterformalin for later anatomic and morphological analysis.The radula of the specimen MNHNCL6640 was dissected,mounted on tape, critical-point dried, coated undervacuum with gold and examined with a JEOL T-300scanning electron microscope (SEM).

Description, measurements and counts followed Roper& Voss (1983). Abbreviations used are the following: TL:total length; DML: dorsal mantle length; MW: mantlewidth; EO: eye diameter; HdL: head length; HdW: headwidth; AL: arm length 1 to 4R/L; WD: web depth A to E;ASC1-4: arm sucker count 1 to 4R/L; AS: arm suckerdiameter; GiLC: gill lamella count; FuL: funnel length; FFL:free funnel length; CaL: calamus length; LL: ligula length;PAL: pallial aperture length; AW: arm base width; GiL:gill length; AF: arm formula; WF: web formula.

The specimens were deposited in the Museo Nacionalde Historia Natural, Chile (MNHNCL), and MuseoZoológico de la Universidad de Concepción (MZUC-UCCC). The comparative material examined is depositedin Zoological Museum of Hamburg (ZMH), Germany.

Figure 1. Map of the coast of Chile

showing capture locations of

Graneledone sp. / Mapa de la costade Chile mostrando los lugares de

captura de Graneledone sp.


Total DNA, to compare with other Graneledonespecies from GenBank, was extracted from 2 specimensfollowing the saline extraction protocol (Aljanabi &Martinez 1997). PCR amplifications were carried out usingfor each sample 0.3 μl of Taq DNA polymerase and 2.5 μlcommercially supplied buffer, with 2 μl dNTPs, and 0.5 μlof each primers LCO1490 and HCO2198 for CytochromeOxidase I (COI) and 16SF and 16SR for 16S rRNA (seeprimers in Allcock et al. 2008). After an initial denaturation(3 min at 94°C), the reaction mixtures were subjected to 35cycles of 94°C (40 s), 50°C (40 s) for COI and 52°C (40 s)for 16S, and 72°C (60 s) followed by a final extension at72°C (7 min) using a thermal cycler. PCR products werepurified by the Wizard™ Prep system (Promega) followingthe manufacturer’s protocols. Purified PCR products weresequenced by Macrogen Inc. Sequences were alignedusing Clustal W implemented in MEGA 5.0 software(Tamura et al. 2011).

Phylogenetic reconstruction and distances betweenspecies were calculated from a matrix including theconcatenated dataset (16S + COI). For this purpose, wedetermined the congruence on the phylogenetic signalof the genes with the ILD test (Farris et al. 1995),implemented in the partition homogeneity test of PAUP*version 4.0b (Swofford 2002). Parsimony (P) and maximumlikelihood (ML) methods, and Bayesian inference (BI) wereapplied to the evaluation of phylogenetic relationshipsof Graneledone species. The evolutionary model thatbest fit the data was d GTR+Γ+I model (AIC = 4067.15,-lnL = 2016.57), determined by the Akaike InformationCriterion (AIC) as implemented by JModelTest (Posada2008). Bayesian analyses were conducted using MrBayesv3.2 (Ronquist et al. 2012) with four default heated chains,each with five million generations, sampled every 1000generations. Runs were checked for convergence see thelikelihood using the Tracer version 1.5 (Rambaut &Drummond 2009). The first 500 trees of each run werediscarded as burn-in, and a consensus of the remainingtrees was computed for the final outcome. Parsimony andML analyses were carried out in PAUP* (Swofford 2002)(heuristic search, tree bisection-reconnection) andsupport for nodes was estimated by bootstrapping with10,000 pseudo-replicates (Felsenstein 1985), providing anestimate of the confidence limits for the resultingtopologies. Finally, we used FigTree 1.3.1 to edit the trees(Rambaut 2009). To construct these trees we usedPareledone charcoti (Joubin, 1905), P. turqueti (Joubin,1905), Megaleledone setebos (Robson, 1932),Thaumeledone peninsulae Allcock et al. 2004, T. rotunda

(Hoyle, 1885) and T. gunteri Robson, 1930 as outgroupsand the phylogenetic tree is rooted using Adelieledonepolymorpha (Robson, 1930) and A. piatkowski Allcocket al. 2003 since these species are basal to Graneledone(Strugnell et al. 2008 a,b). Sequences generated in thisstudy are available from GenBank (Table 1).

SYSTEMATICS

Family Octopodidae d’Orbigny, 1840

Genus Graneledone Joubin, 1918

Graneledone sp. (Figs. 2-6)

DIAGNOSIS

Sucker row uniserial. Ink sac absent. Funnel organ VV-shaped. Hectocotylus clearly differentiated into calamusand ligula; the latter with or without creases. Terminalorgan (penis) coiled in spiral. Gills small, with 5-9 lamellaeper demibranch. Radula homodont or heterodont. Salivaryglands small. Wart-like tubercles cover dorsal surfaces ofmantle, head, arms and web.

Type species: Graneledone verrucosa (Verril, 1881)

Synonymy: Eledone verrucosa Verrill, 1881, Moschitesverrucosa Berry, 1917.

MATERIAL EXAMINED

Adult male 110 mm ML (North off Mocha island, 37º46.17’S,74º07.29’W, southeastern Pacific, 1482 m depth) MNHNCL6641. Collected by J. Sellanes, October 2007. Adult male160 mm ML (off Constitución coast, caught by shrimp

Table 1. Octopod species included in the phylogenetic analyses /

Especies de octópodos incluidas en los análisis filogenéticos


trawling, 35º10’S, 72º55’W, 436 m) MNHNCL 6640.Collected by C.M. Ibáñez, October 2000. Adult female 165mm ML (Northwest off Concepción, caught by a fishingline, 36°15.71’S, 73°43.48’W, 600 m) MZUC-UCCC 32743.Collected by M. Pedraza, March 2007. Adult male 80 mmML (from southern Chile, caught by shrimp trawling)MNHNCL 300039 (without collecting data).

COMPARATIVE MATERIAL EXAMINED

Graneledone antarctica, adult male 55 mm ML(Antarctica, 74°31’54’’S, 027°13’24’’W, 2103 m) ZMH12658, collected during RV Polarstern cruise Antarktis XV/3 at station 134 in 1998. Graneledone antarctica, adultfemale 60 mm ML (Antarctica, 69°25’00’’S, 005°20’00’’W,1800 m) ZMH 72401, collected during RV Polarstern cruiseAntarktis XXIII/2 at station 57 in 2005. Graneledoneverrucosa, two females 46 and 17 mm ML, (NE Atlantic,49°46’00’’N, 012°31’00’’W, 2000 m) ZMH 12657, collectedduring RV Walther Herwig cruise #47 at station 630 in1981. Graneledone verrucosa, adult female 98 mm ML (NAtlantic, 1200 m), ZMH 12663, collected during RVWalther Herwig cruise #46 at station 591 in 1981.Graneledone yamana, adult male 50 mm ML (SE toFalkland Islands, 54°18’00’’S, 56°10’00’’W, 560 m) ZMH12669, collected during RV Walther Herwig cruise #31 atstation 584 in 1978. Graneledone yamana, adult male 58mm ML (near to Falkland Islands, 50°18’00’’S,56°49’00’’W, 515 m) ZMH 2789, collected during RVWalther Herwig cruise #45 at station 325 in 1966.

DIAGNOSIS

Medium sized octopus (TL = 520 to 810 mm) lacking inksac. Suckers small and uniserial (6-8 mm maximumdiameter). Eyes large (20% of DML) and projecting. Funnelorgan VV shaped, free zone of the funnel correspondingto 53-62% of funnel length. First pair of arms alwayslongest (75-78% of TL), with arm formula 1.2.3.4.; webvery deep, sectors unequal; sector A and B always largestand sector E smallest. Third right arm of maleshectocotylized with 43 and 45 suckers and shorter thanopposite arm. The opposite arm with 98 to 107 suckers.Calamus of medium size (50% of the ligula) with deepmedian incision; ligula without copulatory lamellae, butwith 5 to 7 creases. Six to 7 lamellae per demibranch. Radulabearing 7 teeth: 3 central, 2 lateral and 2 marginal plates ineach transversal line. Body surface covered by complexpapillose warts with many tiny spine-like structurescovering body dorsally from mantle to arm tips. Thirty to35 warts at the dorsal mantle midline from anterior to

posterior and 15 to 20 between the eyes. These wartscomposed of 1 to 5 individual processes. Two clusters of18-22 warts above each eye.

DESCRIPTION

Adult specimens examined are of medium size (TL 520 to810 mm). Mantle sacciform (up to 100 mm ML) (Fig. 2A).Body surface covered by complex papillose warts withmany tiny spine-like structures covering body dorsallyfrom mantle to arm tips (Figs. 2A, 5A-B). Thirty to 35warts at the dorsal mantle midline and 15 to 20 betweenthe eyes. These warts are clustered between 1 to 4individual processes in the mantle and webs. Over eacheye, two groups of 18 to 22 individual processes (Fig.5B). These clusters are 1 to 5 mm in diameter. Pallialaperture moderately wide with respect to the mantle width(41-62% of ML). Head width similar to mantle (95 to 100%of MW). Eyes relatively large (diameter = 22 to 32 mm),located in a more lateral than frontal position, almost atthe same level of the dorsal surface. Funnel medium sized(28 to 36% of ML); funnel free portion is very shortrepresents 53 to 62% of the funnel total length. Funnelorgan is ‘VV’ shaped, with internal components widerand longer than external ones (Figs. 2B, 5D).

Arms long and similar in length, generally not exceeding78% of the TL, with the first pair always being the longest,and the fourth pair the shortest. Typical arm formula1.2.3.4. Arms narrow and sharpened to the ends (14 to18% of ML) (Table 2). Web short with E sector shortest.Web formula BCADE, ACDBE and ABCDE (Table 2).

Arms with uniserial suckers (Fig. 2A, 5C). Suckers small,tubular, sessile and without acetabulum aperture (4.2 to5.4% of ML). Third right arm hectocotylized (53 to 48% ofTL), being shorter in length than its opposite (57 to 64%of TL) (Table 2, Fig. 2A-C, 5E). Suckers on thehectocotylized arm number between 43 and 45; oppositearm carries between 98 and 107 (Table 2). Spermatophoriccanal smooth and without pigmentation, extending alongventral face of hectocotylized arm ending in the calamus.Copulatory organ with a small ligula (3.2 to 4.1% ofhectocotylized arm), with 5 to 7 transverse creases (Figs.2C, 5E). Calamus (8 to 9 mm) without pigmentation andslightly long (50% of ligula length). Gills small with six toseven lamellae per demibranch.

Anterior salivary gland small (4 mm); posterior salivaryglands small (6 mm); esophagus slender, pseudocropwithout diverticulum, stomach muscular; caecum smalland striated coiled in a single whorl; large digestive gland


Table 2. Measurements (mm) and counts of Graneledone sp. For abbreviations see Materials and

Methods. *Incomplete or damaged organ / Medidas (mm) y conteos de Graneledone sp. Paraabreviaciones ver Materiales y Métodos. *órgano dañado o incompleto


Figure 2. Graneledone sp. (MNHNCL 6641). (A) adult male, (B) funnel organ,

(C) tip of the hectocotylus. Scale bars: A, 30 mm; B, 30 mm; C, 8 mm /

Graneledone sp. (MNHNCL 6641). (A) macho adulto, (B) órgano del sifón, (C)hectocotilo. Escala de barras: A, 30 mm; B, 30 mm; C, 8 mm

Figure 3. Digestive tract of Graneledone sp. (A) Digestive tract; asg,

anterior salivary glands; eso, oesophagus; cr, crop; sto, stomach;

cae, caecum; bm, buccal mass; psg, posterior salivary glands; dg,digestive gland; pa, pancreas; int, intestine, (B) beaks; r, rostrum;

h, hood; cre, crest; lw, lateral wing; w, wing; (C) radula; p, plate; m,

marginal tooth; ls, second lateral tooth; lf, first lateral tooth; ra,rachidian teeth. Scale bars: A, 10 mm; B, 20 mm; C, 1 mm / Tracto

digestivo de Graneledone sp. (A) Tracto digestivo; asg, glándulas

salivares anteriores; eso, esófago; cr, buche; sto, estómago; cae,ciego; bm, masa bucal; psg, glándulas salivares posteriores; dg,

glándula digestiva; pa, páncreas; int, intestino, (B) picos; r, rostro;

h, capucha; cre, cresta; lw, ala lateral; w, ala; (C) rádula; p, placa;m, diente marginal; ls, diente lateral secundario; lf, diente lateral

primario; ra, diente raquídeo. Escala de barras: A, 10 mm; B, 20

mm; C, 1 mm


Figure 4. Reproductive system of Graneledone

sp. (A) female reproductive system; do, distaloviduct; og, oviductal gland; po, proximal

oviduct; ov, ovary, (B) egg, (C) male reproductive

system; ag, accessory gland; sgl, spermaticgland; tes, testis; ns, Needham’s sac; to,

terminal organ, (D) spermatophore. Scale bars:

A, 10 mm; B, 2 mm; C, 20 mm; D, 15 mm /Sistema reproductivo de Graneledone sp. (A)

sistema reproductivo de hembra; do, oviduct

distal; og, glándula oviductal; po, oviductproximal; ov, ovario, (B) huevo, (C) sistema

reproductivo de macho; ag, glándula

accesoria; sgl, glándula espermática; tes,testículo; ns, saco de Needham; to, órgano

terminal, (D) espermatóforo. Escala de barras:

A, 10 mm; B, 2 mm; C, 20 mm; D, 15 mm

Figure 5. Photographs of Graneledone sp. (MNHNCL 6641). (A) fresh specimen, (B) eye, (C) suckers, (D) funnel organ, (E) hectocotylus, (F)

radulae. Scale bars: A, 30 mm; B, 20 mm; C, 5 mm; D, 20 mm; E, 10 mm; F, 1 mm / Fotografías de Graneledone sp. (MNHNCL 6641). (A) espécimenfresco, (B) ojo, (C) ventosas, (D) órgano del sifón, (E) hectocotilo, (F) rádula. Escala de barras: A, 30 mm; B, 20 mm; C, 5 mm; D, 20 mm; E, 10

mm; F, 1 mm


(43 mm) roundish without ink sac; intestine long (107mm) without anal flaps (Fig. 3A). Upper beak with strongdeep jaw angle; lower beak with distinct groove alonglower edge of insertion plate (Fig. 3B). Radula with amulticuspid rachidian, and to each side two lateral teeth,one marginal tooth and the marginal plate (Fig. 3C, 5F,6A, 6B).

Genitalia of female with large ovary (41 mm diameter);oviductal gland large (11 mm diameter); distal oviductlarge (51 mm length) (Fig. 4A). 55 eggs (10-12 mm long) inovary of maturing female (Fig. 4B).

Male reproductive system with short, thick proximal vasdeferens; spermatophoral gland moderately long (60 mm)(Fig. 4C). Accessory gland large (40 mm), with terminalrecurvature. Spermatophore sac long, containing fourspermatophores (160-180 mm) (Fig. 4D). Poor preservationcondition of the spermatophores did not allow furtherdetailed description. Distal vas deferens short, penisdiverticulum large with spiral, penis large (25 mm) (Fig. 4C).

Locations. Southeastern Pacific Ocean, north of Mochaisland, Chile, 37º46.17’S, 74º07.29’W, at 1482 m (Fig. 1).

Distribution. Southeastern Pacific Ocean, from 35ºS -38º S, 436 to 1482 m depth (Fig. 1).

Habitat. Upper continental slope, associated with hardgrounds (e.g., at methane seep sites).

Table 3. Morphological comparison between Graneledone species / Comparación morfológica entre las especies deGraneledone

Figure 6. Radula of Graneledone sp. (MNHNCL 6640). (A) generalview, (B) detail of the rachidian tooth. Scale bars 0.1 mm / Rádula

de Graneledone sp. (MNHNCL 6640). (A) vista general, (B) detalledel diente raquídeo. Escala de barras 0,1 mm


MORPHOLOGICAL COMPARISON

Some morphological characters distinguish Graneledonesp. from other congeneric species. The presence of supra-ocular cirri in G. boreopacifica, G. verrucosa, G. macrotylaand G. yamana contrasts with the absence of thesestructures in G. sp. Counts of cartilaginous clusters alongthe mantle and head are higher in individuals of G. sp.than in G. gonzalezi, G. macrotyla, G. yamana, G.boreopacifica, G. verrucosa and G. taniwha, but less thanin G. antarctica and G. challengeri (Table 3). The numbersof wart processes within each cluster in G. sp. are higher(1-22 units) and larger (diameter 1-5 mm), than in G.antarctica (1-11 units) (diameter 0.3-0.9 mm), G. gonzalezi(1-12 units) (diameter 3.3 mm), G. boreopacifica (3-12 units)(diameter 1-3 mm), G. verrucosa (1-12 units) (diameter 1-3mm) and G. challengeri (1-10 units) (diameter 1-2.7 mm)and less than G. taniwha (1-37 units) (diameter 0.5-8 mm).Moreover, Graneledone sp. have 5-7 transverse creasesin the ligulae, while G. gonzalezi, G. yamana, G.boreopacifica, G. verrucosa, G. challengeri lacks them.Graneledone taniwha taniwha have 7-8 transversecreases in the ligulae, while G. antarctica have 9-13, highercount than in G. sp., but G. taniwha kubodera have less(3-4) than G. sp. Hectocotylized sucker count is very closebetween G. sp. and G. challengeri, G. verrucosa and G.taniwha, but higher than other species (Table 3).

MOLECULAR PHYLOGENETIC ANALYSIS

Alignment lengths for COI and 16S rRNA fragments were655 bp and 497 bp, respectively. Partition homogeneitytest did not show significant differences between thesemolecular markers (P = 0.21), and sequences werecombined and analyzed as a single data set. The combineddata (16S + COI) of 1152 bp length contained 71 charactersthat were parsimony informative. The Graneledone sp.specimens differed by 9 substitutions (0.8%) from G.boreopacifica, and by 14 substitutions (1.2%) from G.verrucosa and 27 (2.3%) from G. antarctica. ComparingG. boreopacifica with G. verrucosa we found only 17substitutions (1.5%). The only 16S rRNA sequence of G.taniwha from South West Pacific Ocean (GenBankaccession number AJ311119) showed a substantialdivergence (4.2%) with G. sp. Unfortunately, we are notsure if this sequence corresponds to some G. taniwhaspecies because there is no collection informationassociated to the record.

All phylogenetic analyses (P, ML, BI) depict a similartree topology with some slightly differences in nodesupport by bootstrap and posterior probabilities (Fig. 7).

These high nodes values support the hypothesis ofmonophyly of Graneledone (Fig. 7). Graneledoneboreopacifica was the sister species of Graneledone sp.(Fig. 7).

We found a ML tree of -lnL = 1159.72. Maximumparsimony analysis using the combined data set(16S+COI) returned one parsimonious tree, 212 steps inlength (consistency index excluding uninformative sites= 0.761; retention index = 0.802).

DISCUSSION

Morphological, molecular and phylogenetic analysis ofmtDNA sequences of the Chilean Graneledone specimensand those of North Pacific, North Atlantic and Antarcticasupported the taxonomic distinctiveness of thesespecimens as a candidate species. However, furtherrevision of the holotype and/or paratypes of the otherspecies of Graneledone are needed to confirm the identityof Graneledone from Chile. Holotypes of Graneledonespecies are in France, Russia, England, Germany and NewZealand (Sweeney 2001) making them very difficult toaccess and study for Southern Hemisphere researchers.

The great genetic similarity between Graneledonespecies is due to their recent origin, estimated to be about3 million years (Strugnell et al. 2008a). Similar low

Figure 7. Bayesian phylogram of Graneledone species (16S+COI).Node values are bootstrap of 10,000 iterations of MP and ML, and

posterior probabilities of BI / Filograma Bayesiano de las especies

de Graneledone (16S+COI). Valores en los nodos son el resultadode bootstrap de 10.000 iteraciones de P y ML, y probabilidades

a posteriori de BI


differentiation between inter-species comparisons wasdetected in the related octopodid genus Pareledone (1-2%; Allcock et al. 2007) and Thaumeledone (2-3.3%;Strugnell et al. 2008b). The phylogenetic relationship isvery consistent with the hypothesized monophyly ofGraneledone (Voight 2000a) and the Antarctic origin ofthese deep-sea octopuses (Strugnell et al. 2008a), becauseG. antarctica is the most basal species of Graneledoneand the Atlantic and Pacific species are the most derivatedand the divergence of Pacific species could be veryrecent. Sequences of all Graneledone species are neededto complete the phylogenetic reconstruction, establishtheir origin and the divergence of these deep-sea species.

The great variation in morphometric and meristic datain species of the genus Graneledone (Table 3) complicatesthe identification, given the overlap in counts of gills,suckers and warts among different species. Multivariateanalysis of the morphological dataset suggests that thesecounts and indices, traditionally used for discriminatingbetween cephalopods species, do not show greatdiscrimination at species level, but provide excellentdiscrimination at the generic level (Allcock et al. 2008).The difficulty differentiating between Graneledone sp.and their relatives is consistent with the idea of crypticspecies, and as mentioned earlier, could be a consequenceof the recent origin of the group. Voight (2000a)commented on the possible existence of cryptic speciesof Graneledone within the north Pacific specimenscollected at different depths. Cryptic speciation inoctopuses has been reported in other genera like Octopusand Pareledone (Söller et al. 2000, Allcock et al. 2011),and could be more common than previously thought.

The records of octopus with uniserial suckers fromChile had many problems. Roper et al. (1984) mentionedPareledone spp. with circumpolar distributions includingthe coast of southern Chile, Argentina and South Africa,leading Rocha (1997) to include Pareledone charcoti andP. turqueti in the check list of cephalopods from Chile.Guerrero-Kommritz (2006) reviewed the octopods withuniserial suckers from southwest Atlantic and finds a verydiverse octopus fauna including Eledone, Graneledone,Vosseledone, Thaumeledone, and Pareledone. However,we studied up to 2500 km of Chilean coast and the onlyoctopuses with uniserial suckers were Graneledone (seeIbáñez et al. 2011). Thore (1959) reported a specimen ofBentheledone rotunda (Hoyle 1885) (now Thaumeledonerotunda) off Valparaíso, Chile (33°S, 71°W), but hementioned that the identification must be considereddubious, since H.M.S. Challenger only captured the arms

of the octopus. This specimen could have correspondedindeed to Graneledone sp., since at this latitude thisspecies is so far the only reported deep-sea octopus withone row of suckers, hence, T. rotunda is not distributedin Chilean waters as has been reported (Rocha 1997, Vegaet al. 2001, Vega 2009). Furthermore, the specimenidentified as G. antarctica (MNHNCL 300039) by Vega etal. (2001) from southern Chile corresponds to Graneledonesp. (Table 1).

Although some of the specimens have been foundassociated with hard grounds, like those present atmethane seeps, no special adaptations to this type ofhabitat have been observed. We assume that thepreference of this species for these sites is just aconsequence of the locally enhanced abundance ofpotential prey and the availability of hard substrategenerated by the carbonate reefs present at seep sites(Sellanes et al. 2008).

ACKNOWLEDGMENTS

We thank Janet Voight, Jan Strugnell and Jürgen Guerrero-Kommritz for their valuable comments of the early versionof this manuscript. We are deeply indebted to captainand crew of AGOR Vidal Gormáz of the Chilean Navy, aswell as scientific staff during VG07 cruise for their supportat sea. We thank Sergio Letelier and Bernhard Hausdorffor help in examining specimens from the MNHNCL(Chile) and ZMH (Germany), and Milton Pedraza forcollecting one specimen. FINANCIAL SUPPORT: Thiswork was partially funded by grants to C.I. FONDECYT3110152, to J.S., FONDECYT 1061217 and FONDECYT1100166, and to E.P., ICM P05-002 and PFB-23. Support toM.C. Pardo-Gandarillas by a MECESUP-Chile DoctoralFellowship are also acknowledged.

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Received 20 June 2012 and accepted 4 October 2012

Associate Editor: Mauricio Landaeta D.

doi 10.4067/s0718-19572012000300007 article morphological … · 2013-07-30 · key words:...

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